Differential Regulation of Two 3′ End Variants of P450 Aromatase Transcripts and of a New Truncated Aromatase Protein in Rabbit Preovulatory Granulosa Cells
Résumé
In rabbit granulosa cells, two cytochrome P450 aromatase (P450 arom) mRNAs issued from promoter II were described: a full-length and a truncated transcript. Western blot analysis showed two P450 arom proteins with apparent molecular masses of 53 and 46 kDa, which are consistent with the predicted theoretical sizes of proteins encoded by these two transcripts. To examine the involvement of the truncated transcript in the regulation of P450 arom gene expression, the level of each transcript was specifically quantified in cultured granulosa cells by competitive quantitative RT-PCR. FSH induced a dose-dependent increase in both estradiol production and P450 arom mRNAs levels with a much more enhancement in the full-length mRNA. The half-life of the transcripts could not explain this differential regulation. Upon dibutyryl cAMP stimulation, the full-length mRNA was less abundant than the truncated one. In contrast, Western blot analysis revealed a stimulation of the 53-kDa protein content, whereas the 46-kDa protein amount was apparently unaffected. TGF beta in FSH-stimulated conditions decreased both estradiol production and P450 arom transcripts levels. TGF beta did not modify estradiol production and aromatase protein amounts induced by dibutyryl cAMP, whereas the two P450 arom mRNAs levels were increased. In conclusion, we report for the first time that a protein encoded by a truncated P450 arom mRNA could be involved in the regulation of estrogen production. Moreover, we show that the two P450 arom mRNAs are regulated in a differential manner, probably through hormonal control of the alternative splicing.